Primary human macrophages from healthy individuals show differential susceptibility and inflammatory response to Mycobacterium avium

Title: Primary human macrophages from healthy individuals show differential susceptibility and inflammatory response to Mycobacterium avium
Author(s): T Shaw A Krasnodembskaya G Schroeder C O'Kane
Institution: Queen's University Belfast
Poster: Click to view poster
Category: General Respiratory and Sleep
Abstract: Non-tuberculous mycobacterial pulmonary disease (NTM-PD) is increasingly reported in patients with no identifiable risk factors. Moreover, inter-strain heterogenicity in infectivity and virulence in NTM has been reported in macrophage cell lines, but not in primary macrophages. We sought to characterise the susceptibility of primary macrophages from healthy individuals to Mycobacterium avium (M. avium).

Primary monocyte-derived macrophages (MDMs) from healthy volunteers were infected with M. avium (Chester laboratory reference strain, 104 clinical reference strain and a clinical isolate from the Northern Ireland Mycobacterial Reference Laboratory) at multiplicity of infection of 0.1-1 for 4 hours, before washing. Supernatant was collected, and cells lysed in 0.2% saponin, to determine extracellular and cell-associated colony counts at baseline and 24-hour intervals. Supernatant cytokines were quantified by ELISA. Statistical analysis was performed using the Kruskal-Wallis test.

There was marked heterogenicity between MDMs from 22 healthy donors in susceptibility and inflammatory response to infection with M. avium Chester strain. Bacterial CFU rose from baseline by median 1.7-fold after 24 hours (IQR 1.2-3.1) and 3.1-fold after 72 hours (IQR 2.3-5.2). Some outliers exhibited control of bacterial growth up to 72 hours, whereas others acceded to 10-fold growth in CFU counts.

Concentrations of pro-inflammatory cytokines in the supernatants of infected MDMs after 72 hours varied between donors. The greatest heterogenicity was noted in IL-6 concentration (n=12, median 1574, IQR 477-5110 pg/ml) followed by IL-8 (n=11, median 16956, IQR 9320-18356 pg/ml) and TNF-α (n=13, median 908, IQR 481-1306 pg/ml).

Differential infectivity and proliferation was observed between strains of M. avium. After 4-hour incubation with MDMs, the median percentage of inoculating CFU that cell-associated was higher in the Chester strain (25.4%) than the 104 strain (5.7%, p<0.01) and CI5 strain (10.0%, p=0.052). There was no significant difference in the rate of proliferation over 72 hours between the Chester and CI5 strains (median 3.1-fold and 3.0-fold respectively). However the 104 strain total CFU count decreased in nearly all donors (median 0.5-fold, IQR 0.3-0.8-fold) suggesting this strain displays less virulence in intracellular infection.

In conclusion, primary macrophages from healthy donors show variable susceptibility and inflammatory responses to different strains of M. avium. Further investigation in mycobacterial host-pathogen interactions is warranted to predict susceptibility to NTM-PD and identify antimicrobial strategies.